Fig. 2

Impact of LIN7A gene silencing on U87 cell proliferation, invasion, and clone formation. Impact of LIN7A silencing on U87 cell growth. The RTCA proliferation assay was performed by seeding cells onto E-plate plates at initial densities of 2500 cells (A) and 4000 cells (B). Cell proliferation was monitored over 24 h, and the growth curves of U87shLIN7A (green) and U87Ctrl (red) were compared. Statistical analysis indicated a significant difference between the two groups (P < 0.05). LIN7A silencing enhances U87 cell invasion. The RTCA invasion assay (C) was performed using CIM-plate plates to evaluate and compare the invasion profiles of U87shLIN7A (green) and U87Ctrl (red) cells. The analysis revealed a statistically significant difference between the two invasion curves (P < 0.05).For the spheroid invasion assay (D), 3D invasion of U87 spheroids into the brain-mimicking matrix (BMM) was monitored over time. Phase contrast microscopy images were captured at 0, 24, 48, and 72 h of culture. The invasion areas were quantified using ImageJ software, and the data showed statistically significant differences between the two groups (E) (*P < 0.05). LIN7A gene silencing inhibits clonogenicity of U87 cells clonogenic assay (F): cells were seeded at varying concentrations in each well, cultured under standard conditions, and monitored for colony formation over a 14-day period. The number of colonies formed for each initial cell density was quantified (as shown in Supplementary Figure S1). Statistical analysis revealed a significant difference between the two groups, with U87Ctrl cells requiring a minimum of 4.48 cells to form colonies, compared to 52.38 cells for U87shLIN7A cells (P < 0.01).